This invention relates to a process for producing deacetylase by incubating a deacetylase-producing bacterium belonging to Vibrio sp.
The term "deacetylase" as used herein means an enzyme capable of deacetylating N-acetylglucosamine, oligomers thereof or chitin.
In recent years, chitin, which is a biomass resource following cellulose, and chitosan, which is a deacetylation product thereof, have attracted public attention as polysaccharides having a number of useful functions. It has been found out, in particular, that chitosan of a low molecular weight (i.e., a glucosamine oligomer) has potent antimicrobial and antitumor activities per se and, furthermore, serves as an elicitor capable of inducing the production of a substance having an antimicrobial activity.
Compared with chitin, chitosan homologs are highly soluble and thus have become more and more important as a starting material for producing a number of useful substances.
Chitosan has been produced by removing inorganic salts, proteins and lipids from the cuticle (crust) of a crustacean such as prawn or crab and heating the purified chitin thus obtained in a concentrated alkali solution of 30 to 60%. Namely, chitosan is deacetylated chitin.
When heated in the aforesaid concentrated alkali solution, however, the chitin undergoes decomposition of the main chitin chain simultaneously with the deacetylation, which causes such a problem that the yield of the target chitosan is lowered while the amounts of by-products are increased.
There has been known a biological method for deacetylating chitin with the use of deacetylase or a microorganism capable of producing this enzyme.
There has been reported no example of this biological method, except one wherein a microorganism of Mucor sp. is used, one wherein a microorganism of Aeromonas sp. is used and one wherein a microorganism of Colletotrichum sp. is used.
With the use of the microorganism of Mucor sp., soluble glycol chitin and the pentamer can be deacetylated to the extent of 12 to 14%. However other oligosaccharides and solid chitin can be deacetylated thereby to the extent of as low as 0 to 5% [refer to Y. Araki, E. Ito, Eur. J. Biochem., 55, 71 (1975)].
The microorganism of Aeromonas sp. is a strain searched for in order to treat exclusively solid chitin of a high degree of polymerization. Therefore nothing is reported relating to the effects of this microorganism on oligosaccharides [refer to K. Shimabara, K. Iwasaki, Reports of the Asahi Glass Foundation for Industrial Technology, 41, 299 (1982)].
The microorganism of Colletotrichum sp., which suffers from a decrease in the activity at 30.degree. C., should be incubated at a relatively low temperature (e.g., 25.degree. C.). Thus the deacetylation with the use of this microorganism requires a long period of time (about 5 days). In addition, it has only a low deacetylation activity on N-acetylglucosamine trimer or dimer. Namely, the ratios of its deacetylation activity on the trimer and the dimer, based on that on the pentamer, are respectively 6% and 1% [refer to H. Kauss, B. Bauch, Methods in Enzymology, 161, 518 (1988)].